scholarly journals Targeted Proteomics of Pulmonary Hypertension in Sickle Cell Disease

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 981-981
Author(s):  
Varshini Babu ◽  
Jane A. Little ◽  
Claudia R. Morris ◽  
Roberto Machado ◽  
J Simon R Gibbs ◽  
...  
Keyword(s):  
Pulmonary Hypertension ◽  
Sickle Cell Disease ◽  
Sickle Cell ◽  
Negative Correlation ◽  
Neutrophil Count ◽  
Tricuspid Regurgitation ◽  
Research Funding ◽  
Cell Disease ◽  
Protein Biomarkers ◽  
False Discovery

Abstract BACKGROUND AND AIM: Hemolysis, inflammation and coagulopathy associated with sickle cell disease (SCD) can lead to pulmonary hypertension. More than 15% of adult patients with SCD are affected by pulmonary hypertension as measured by a tricuspid regurgitation velocity (TRV) ≥ 2.9 m/sec (Nouraie, M. et al., Validation of a composite vascular high-risk profile for adult patients with sickle cell disease. American Journal of Hematology, 94:E312-E314. 2019). Moreover, sickle cell disease patients with pulmonary hypertension have a much higher mortality risk than those without pulmonary hypertension. There is little knowledge on the proteomic profile of pulmonary hypertension in SCD. Our aim was to conduct a proof-of-concept study to explore targeted serum protein biomarkers that are differentially expressed in SCD patients with elevated tricuspid regurgitation velocities and explore their corresponding pathways. MATERIAL AND METHODS: Data from the Walk-PHaSST study was used for this study. The Walk-PHaSST was a multicenter study to assess the effect of sildenafil for SCD patients with pulmonary hypertension (Gladwin, M. T. et al., Risk factors for death in 632 patients with sickle cell disease in the United States and United Kingdom. PLoS One, 9:e99489. 2014). In the screening phase of the study, 720 patients with SCD were recruited. Study participants underwent clinical and lab examinations, as well as an echocardiography. We selected two groups of patients: one group of patients with a TRV≤2.6 m/sec and another with TRV≥2.9 m/sec (N =35 in each). The serum concentration of 92 protein biomarkers was measured using an OLINK cardiovascular panel. This panel assess a range of biological process including inflammatory response, angiogenesis, cell adhesion, coagulation, and response to hypoxia. T-tests were performed between the aforementioned groups for each of the serum biomarkers. The Hochberg method was used to calculate the false discovery rate. A volcano plot was created using the Bonferroni correction. Finally, we tested the correlation of the differentially expressed biomarkers with clinical variables measured in blood samples of the participants. These variables include white blood cell (WBC), neutrophil count, thrombospondin-1 (TSP as measure of coagulation), PIGF and VEGF (growth factors involved in angiogenesis), and NT-proBNP (elevated in pulmonary hypertension). RESULTS: Using a false discovery rate of 0.01, we discovered 14 significantly expressed proteins. Six of them passed a Bonferroni corrected overall critical p value < 0.00054 (Figure 1). These included T-cell surface glycoprotein (CD4), lymphotactin (XCL1), SLAM family member 7 (SLAMF7), galectin-9 (GAL9), TNF-related apoptosis-inducing ligand receptor 2 (TRAILR2), and tumor necrosis factor receptor superfamily member 11A (TNFRSF11A). We observed up to a 1.2-fold increase in these 6 protein biomarker levels in the high TRV groups. The correlogram (Figure 2) indicated that there is a slightly positive correlation between WBC and CD4, GAL9, TRAILR2, and TNFRSF11A (r > 0.20). For neutrophil count, we observed significantly negative correlation with selected markers including TNFRSF11A levels (r = -0.51), GAL9 (r = -0.35), XCL1 (r = -0.27), SLAMF7 (r = - 0.21), and CD4 (r < - 0.24). A significantly negative correlation between TSP and GAL9 levels (r = -0.31) was also observed. Finally, we observed a strong, positive correlation between all proteins and serum NT-proBNP levels (r > 0.44). CONCLUSION: Circulatory protein markers of immune response and coagulation are highly expressed in SCD patients with elevated TRV. This provides evidence that these protein biomarkers have potential to be utilized as prognostic markers for pulmonary hypertension in patients with SCD. Figure 1 Figure 1. Disclosures Little: Biochip Labs: Patents & Royalties; Hemex Health, Inc.: Patents & Royalties. Gibbs: Acceleron Pharma: Consultancy, Other: lecture fees; Actelion: Consultancy, Other: lecture fees; Aerovate: Consultancy, Other: lecture fees; Bayer: Consultancy, Other: lecture fees; Compexia: Consultancy, Other: lecture fees; Janssen: Consultancy, Other: lecture fees; MSD: Consultancy, Other: lecture fees ; Pfizer: Consultancy, Other: lecture fees; United Therapeutics: Consultancy, Other: lecture fees. Gordeuk: Modus Therapeutics: Consultancy; Novartis: Research Funding; Incyte: Research Funding; Emmaus: Consultancy, Research Funding; Global Blood Therapeutics: Consultancy, Research Funding; CSL Behring: Consultancy. Nouraie: Phoenicia BioScience Inc.: Consultancy.

Safety and Efficacy of Sildenafil Therapy for Doppler-Defined Pulmonary Hypertension in Patients with Sickle Cell Disease: Preliminary Results of the Walk-PHaSST Clinical Trial.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 571-571 ◽  
Author(s):  
Roberto F Machado ◽  
Robyn J Barst ◽  
Nancy A Yovetich ◽  
Kathryn L Hassell ◽  
Jonathan C. Goldsmith ◽  
...  
Keyword(s):  
Pulmonary Hypertension ◽  
Sickle Cell Disease ◽  
Adverse Events ◽  
Arterial Pressure ◽  
Sickle Cell ◽  
Research Funding ◽  
Cell Disease ◽  
Safety And Efficacy ◽  
Preliminary Results ◽  
Significant Difference

Abstract Abstract 571 Background: Pulmonary hypertension (PH) is associated with increased mortality in patients with sickle cell disease (SCD). Methods: Walk-PHaSST (treatment of Pulmonary Hypertension and Sickle cell disease with Sildenafil Therapy) is a multi-center (10 United States and United Kingdom Centers), placebo-controlled, double-blind 16-week trial evaluating the safety and efficacy of oral sildenafil for the treatment of Doppler-defined PH (tricuspid regurgitant jet velocity [TRV] ≥2.7m/s) in adults and children (aged >12 years) with SCD. The primary endpoint was the six-minute walk distance (6MWD). The study was designed with a planned screening of approximately 1000 subjects, to enroll 132 subjects for inclusion in the nested Main Interventional Trial (MIT). In the screening trial, subjects were evaluated by history and physical examination, laboratory screening, transthoracic Doppler echocardiography and 6MWD. Randomized subjects were stratified by TRV (2.7-2.9 m/s and ≥3.0 m/s), and those in upper strata underwent a right heart catheterization (RHC). Preliminary Results: Of the 722 screened subjects, 150 (26%) had both a TRV ≥2.7 m/s AND 6MWD of 150-500 meters (m), qualifying for MIT enrollment. A total of 74 subjects (13%) were randomized into the MIT. The study was prematurely stopped due to a statistically significant increase in serious adverse events (SAEs) in the sildenafil arm after 33 subjects had completed the 16 week assessments and 74 subjects (37 sildenafil: 23 female, 47 ± 12 years, TRV 3.0 ± 0.4 m/s, 6MWD 378 ± 93 m; 37 placebo: 23 female, 44 ± 14 years, TRV 2.9 ± 0.3 m/s, 6MWD 381 ± 75 m) had been randomized in the MIT. To evaluate safety and efficacy, all 74 subjects were evaluated, with pre-defined primary and secondary endpoint analysis and imputation rules for missing data. Baseline gender, hemoglobin phenotype, TRV and 6MWD were similar between sildenafil and placebo (all p>0.05). There was a significant increase in SAEs in the sildenafil arm (46% vs. 22% of randomized subjects; p=0.048) but no significant difference in adverse events (AEs; 76% vs. 68%; p=0.607). Sickle cell anemia with crisis (hospitalization defining the SAE) accounted for the significant difference in SAEs (35% vs. 11%; p=0.025). In reference to AEs, patients on sildenafil tended to have more headache (27% vs. 14%; p=0.247) and more blurred vision (11% vs. 3%; p=0.358). No other SAEs or AEs by organ system or preferred term were significantly different (all p > 0.43). There were no AEs classified as life-threatening and there was one death in the placebo arm. To assess potential efficacy, all 33 subjects with TRV of ≥3.0 m/s underwent RHC and received a single test dose of 60 mg of sildenafil; data are currently availabel for 22 of those subjects. Although this dose of sildenafil acutely decreased mean pulmonary arterial pressure (p=0.01), and mean systemic arterial pressure (p<0.01), the change in pulmonary vascular resistance was not significant. There were no apparent safety issues with the acute sildenafil dosing (e.g. priapism). After 16 weeks of sildenafil, there was no difference in the change in TRV (adjusted mean change from baseline: sildenafil -0.10 ± 0.08 m/s, placebo -0.13 ± 0.8 m/s; p=0.7) or in 6MWD (adjusted mean change from baseline: sildenafil -17 ± 20.9 m, placebo +1.4 ± 21.8 m; p=0.47). On the Brief Pain Inventory (BPI), sildenafil subjects reported worsening pain during walking (p=0.07; p=0.17) and less enjoyment of life (p=0.09; p=0.04) vs. placebo at the interim visits (6 and 10 week, respectively). For subjects in the sildenafil group, no difference was detected in 6MWD for those experiencing a VOC vs. those without a VOC (adjusted mean 386 ± 15.9 m; 372 ± 11.7 m; p=0.39). Preliminary Conclusions: In conclusion, sildenafil significantly increased rates of VOCs requiring hospitalization vs. placebo. The premature study termination for safety concerns limited the sample size for efficacy assessments; however, further investigation may be warranted in a more select group of patients with optimized hydroxyurea and transfusion therapy. Based on the completed analyses, no relationship has been established between experiencing pain (via serious VOC) and 6MWD. Additional analyses will focus on determining whether pain (as measured by BPI) is correlated with change in 6MWD. Finally, these data suggest a potential role for the cyclic cGMP axis in the pathobiology of VOC and sickle cell disease related pain. Disclosures: Barst: Pfizer: Consultancy, Research Funding. Gibbs:Pfizer: Honoraria, Membership on an entity's Board of Directors or advisory committees. Girgis:Pfizer: Research Funding. Badesch:Pfizer: Consultancy, Research Funding.

Risk Factors for Death in 632 Patients with Sickle Cell Anemia in the United States and United Kingdom

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 3240-3240 ◽  
Author(s):  
Mark T Gladwin ◽  
Robyn J Barst ◽  
J. Simon R. Gibbs ◽  
Marianna Hildesheim ◽  
Vandana Sachdev ◽  
...  
Keyword(s):  
Risk Factors ◽  
Pulmonary Hypertension ◽  
Sickle Cell Disease ◽  
Sickle Cell ◽  
Research Funding ◽  
Proportional Hazards ◽  
The United States ◽  
Right Heart Catheterization ◽  
Heart Catheterization ◽  
Cell Disease

Abstract Abstract 3240 The role of pulmonary hypertension as a common and attributable cause of mortality in patients with sickle cell disease remains controversial. To assess this question and explore risk factors for death in patients with sickle cell disease we evaluated 632 patients in the Walk-PHASST pulmonary hypertension screening cohort, recruited from nine different study sites in the United States and one site in the United Kingdom. Methods: Patient characteristics and their associations with mortality were analyzed with Cox proportional hazards regression analysis. Based on data from three right heart catheterization screenings studies that have recently been published, we defined the presence of pulmonary hypertension for this analysis by a Doppler-echocardiographic measurement of the tricuspid regurgitant jet velocity (TRV) ≥ 3.0 m/s, which has a 67–75% positive predictive value for a mean pulmonary artery pressure ≥ 25 mm Hg by right heart catheterization. This therefore represents a very conservative threshold for a large population screening study. Among subjects with a measurable TRV (n=572), 64 (11.2%) had measurements of ≥ 3.0 m/sec. Among those with measurable NT-proBNP (n=582), 140 (24.1%) had measurements ≥160 pg/mL, a value associated with both pulmonary hypertension and mortality. A total of 39 (7.4%) had both high TRV (≥3.0 m/sec) and high NT-proBNP (≥160 pg/mL). Results: Over a median follow-up time of 29 months, we observed 22 deaths. 50% (N=11) of these patients had a TRV≥ 3.0 m/sec. At 24 months the cumulative survival was 83% for patients with TRV ≥ 3.0 m/sec and 98% for patients with TRV < 3.0 m/sec (p<0.0001). The unadjusted hazard ratios for death were 11.14 (95% CI 4.1–30.1; p<0.0001) for patients with TRV above and below 3.0 m/sec and 4.55 (95% CI 1.8–11.3; p=0.001) for patients with NT-proBNP above and below 160 pg/mL. For patients with both high TRV (≥ 3.0 m/sec) and high NT-proBNP (≥ 160 pg/mL), the unadjusted hazard ratio was 14.86 (95% CI 5.5–39.9; p<0.0001). Overall, an increased risk of death was observed for both age and gender, with males at higher risk relative to females (HR=2.48, 95% CI 1.0–6.1; p=0.05), and patients older than 47 years (HR=2.02, 95% CI 1.1–3.8; p=0.03). Associations with mortality were also observed for chronic transfusions (HR=3.00, 95% CI 1.2–7.8; p=0.02) and a NYHA/WHO class value or III or IV (HR=4.52, 95% CI 1.4–14.3; p=0.01). Other variables associated with mortality in our cohort included a high hemolytic component, aspartate aminotransferase (AST), ferritin, and creatinine. Variables not associated with mortality included current hydroxyurea use, SC disease, self-reported history of painful episodes, and six-minute walk distance. In stepwise multiple proportional hazards regression analysis, the association between TRV and mortality remained significant after adjustment for all other risk factors, including ferritin, AST, creatinine and even NT-proBNP (HR 4.27, 95%CI 1.3–14.1; p=0.04). Conclusions: Using a more conservative cut-off value of TRV ≥ 3.0 m/sec as defining PH in a large screening population of sickle cell disease patients, PH occurs in approximately 10% of unselected screened patients and is associated with the highest unadjusted and adjusted risk for death of any measured variable. Disclosures: Gladwin: Bayer Corp: Consultancy, Research Funding; NIH and NHLBI: Research Funding; Gilead Sciences: Research Funding. Barst:Ventripoint: Stock options Other; Actelion, Eli Lilly, Gilead, Glaxo Smith-Kline (GSK), Medtronics, Bayer, Ikaria, Pfizer, Novartis, VentriPoint: Consultancy, Honoraria. Girgis:NIH/NHLBI: Research Funding, Travel support Other. Rosenzweig:NIH: Research Funding. Badesch:NIH: Research Funding. Lanzkron:NIH: Research Funding.

PDE-4 Inhibitor Rolipram Prevents Hypoxia Induced Pulmonary Hypertension in Transgenic Sickle Cell Sad Mice.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 3577-3577
Author(s):  
Lucia De Franceschi ◽  
Giorgio Malpeli ◽  
Anne Janin ◽  
Aldo Scarpa ◽  
Christhophe Leboeuf ◽  
...  
Keyword(s):  
Gene Expression ◽  
Pulmonary Hypertension ◽  
Sickle Cell Disease ◽  
Lung Injury ◽  
Sickle Cell ◽  
Neutrophil Count ◽  
Cell Disease ◽  
Endothelin 1 ◽  
Tnf Α ◽  
Peripheral Neutrophil

Abstract Pulmonary hypertension (PH) is one of the major causes of morbidity and mortality for young-adult patients with sickle cell disease (SCD). Little is known about the role of vaso-active agents, pro-inflammatory and pro-fibrotic mediators in the genesis of PH in SCD. We developed a model for studying PH in transgenic sickle cell SAD mice. Wild-type (WT) and SAD mice aged between 4 and 6 months were divided in groups of 7 animals each. One group from each strain served as control under normoxia, while a second group from each strain was exposed to 7 days hypoxia (8% oxygen). The following parameters were evaluated in normoxic and hypoxic groups: lung histopathology, complete blood count, reticulocytes, bronchoalveolar lavage (BAL) total leokocyte and neutrophil count, BAL levels of IL-6, IL-10, IL-1β and TNF-α. The expression of nine genes was examined by quantitative RT-PCR. These included endothelin-1 (ET-1), endothelin-1 receptor (ET-1R), cycloxygenase-2 (COX2), NF-kBp65, transforming-growth-factor-1β (TGF-1β), metalloproteinase-2 and 9 (MMP-2, MMP-9), IL-10 and IL-1β. Two of the seven SAD mice exposed to hypoxia died at day fourth. The remaining five mice were sacrified at day seven and showed: a) increased pulmonary arterial wall thickness and α-actin staining, suggesting a vascular remodeling; b) lung injury compatible with PH in SCD; c) increased BAL total leukocyte and neutrophil count; d) increased BAL IL1β, IL10, IL6 and TNF-α; e) increased peripheral neutrophil and reticulocyte counts, hematocrit and hemoglobin levels. In lungs of hypoxic SAD mice, ET-1, ET-1R, TGF-1β, MMP-2, MMP-9, IL-1β and IL-10 gene expression were induced, with no significant changes in NF-Kbp65 gene expression. These data were consistent with a model of PH in SCD. We then assessed the effect of the PDE-4 inhibitor ROLIPRAM on the PH in seven hypoxic SAD mice. ROLIPRAM was administrated at the dosage of 30 mg/Kg/day by gavage. No deaths were observed in this group. ROLIPRAM a) protected SAD mice from hypoxia induced lung injury; b) decreased BAL total leukocyte and neutrophil count; c) decreased BAL IL-1β, IL-6 and IL-10 levels; d) reduced peripheral neutrophil count; d) normalized and/or modulated the expression of the hypoxia-induced genes. These data suggest that the PDE-4 inhibitor ROLIPRAM has anti-inflammatory properties and could have beneficial effects on the hypoxia-induced airway-remodeling and possibly PH in sickle cell disease.

NT-Probnp as a Marker of Cardiopulmonary Compromise and Exercise Limitation In Adults with Sickle Cell Anemia In the Walk-PHaSST Study

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 1639-1639
Author(s):  
Mehdi Nouraie ◽  
Robyn J. Barst ◽  
Erika Berman Rosenzweig ◽  
Vandana Sachdev ◽  
Roberto F. Machado ◽  
...  
Keyword(s):  
Pulmonary Hypertension ◽  
Sickle Cell Disease ◽  
Brain Natriuretic Peptide ◽  
Natriuretic Peptide ◽  
Sickle Cell ◽  
Sickle Cell Anemia ◽  
Research Funding ◽  
Lateral Wall ◽  
Left Ventricular ◽  
Cell Disease

Abstract Abstract 1639 Background: The N-terminal (NT) pro-brain natriuretic peptide (proBNP) hormone mediates natriuresis and vasodilation and down-regulates the renin-angiotensin-aldosterone axis. Elevated NT-proBNP levels have been reported in left heart failure [1] and pulmonary hypertension, [2] and the degree of NT-proBNP elevation appears to reflect the clinical and hemodynamic status of these patients. [3] Levels ≥160 pg/ml have been associated with pulmonary hypertension in adults with sickle cell disease, with a positive predictive value of 78%. [4] Elevated levels are highly predictive of mortality in the Cooperative Study of Sickle Cell Disease cohort (Machado et al, unpublished observations, 2009). Based on these data, we investigated the relationship of serum NT-proBNP levels with Doppler-echocardiographic estimates of right ventricular systolic pressure and left ventricular diastolic dysfunction, and exercise capacity in a large, prospective, multi-center, international cohort of sickle cell anemia patients in a cross-sectional manner. Methods: Walk-PHaSST (treatment of Pulmonary Hypertension and Sickle cell disease with Sildenafil Therapy) includes an on-going observational study of sickle cell disease patients at nine United States Centers and one United Kingdom Center. In the screening phase of the study, patients had clinical evaluation, echocardiography, six-minute walk (6MW) testing and serum NT-proBNP measured. Of 720 patients screened, 483 had hemoglobin SS phenotype. The study prospectively defined 3 TRV cohorts: < 2.7 m/sec, 2.7–2.9 m/sec and TRV as ≥3.0 m/sec. Results: Of 483 hemoglobin SS patients, the median age was 35 years (range of 12 to 69 years) and the gender split was 250 females to 233 males. The 6MW test was performed in 475 patients, TRV in 453, NT-proBNP concentration in 447, and lateral wall E/Ea in 436. TRV was 2.7–2.9 m/sec in 22% and ≥3.0 m/sec in 17%. The median NT-proBNP concentration was 79 pg/ml with an interquartile range of 35–180. 119 (27%) of participants had levels ≥160 pg/ml. By multiple linear regression, NT-ProBNP (natural log) correlated with creatinine >1.4 mgdL (standardized beta = 0.31; P <0.0005), lower hemoglobin concentration (standardized beta = -0.29; P <0.0005), older age (standardized beta = 0.22; P <0.0005), higher TRV categories (standardized beta = 0.12; P = 0.005), and higher log LV lateral wall E/Ea ratios (standardized beta = 0.08; P = 0.066). NT-proBNP ≥160 had a positive predictive value of 58% for TRV 2.7–2.9 m/sec, 37% for TRV ≥3.0 m/sec, 68% for LV lateral wall E/Ea above the median (>6.4), and 38% for an upper quartile value of LV lateral wall E/Ea (>8.0). By multiple linear regression, higher 6MW distance (6MWD) correlated with lower NT-proBNP levels after adjustment for age and gender. An elevated NT-proBNP level was associated with an estimated 22 m lower 6MWD (P = 0.026). Conclusions: In this prospective, multicenter, international study of patients with sickle cell anemia, elevated NT-proBNP levels appear to be an independent predictor of: 1) higher estimated right ventricular systolic pressure and left ventricular filling pressure as assessed by Doppler-echocardiography and 2) lower 6MWD. These data validate the use of NT-proBNP in screening patients with sickle cell anemia for cardiac dysfunction and exercise limitations, and as an exploratory endpoint in controlled studies. References: 1. Tsutamoto, T., et al., Attenuation of compensation of endogenous cardiac natriuretic peptide system in chronic heart failure: prognostic role of plasma brain natriuretic peptide concentration in patients with chronic symptomatic left ventricular dysfunction. Circulation, 1997. 96(2): p. 509-16. 2. Kragelund, C., et al., N-terminal pro-B-type natriuretic peptide and long-term mortality in stable coronary heart disease. N Engl J Med, 2005. 352(7): p. 666-75. 3. Leuchte, H.H., et al., Clinical significance of brain natriuretic peptide in primary pulmonary hypertension. J Am Coll Cardiol, 2004. 43(5): p. 764-70. 4. Machado, R.F., et al., N-terminal pro-brain natriuretic peptide levels and risk of death in sickle cell disease. JAMA, 2006. 296(3): p. 310-8. Disclosures: Barst: Pfizer: Consultancy, Research Funding. Rosenzweig:Pfizer: Research Funding. Hassell:Novartis: Research Funding. Gibbs:Pfizer: Speakers Bureau. Badesch:Pfizer: Honoraria, Research Funding.

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